Spinal muscular atrophy (SMA) is an autosomal recessive form of muscular dystrophy that afflicts approximately 1 in 6,000 live births and clinically manifests as progressive muscle weakness due to deterioration of motor neurons. SMA results from insufficient levels of the survival motor neuron (SMN) protein, usually due to homozygous deletion of the SMN1 gene. A nearly identical copy gene, SMN2, fails to protect from development of SMA because its mRNA undergoes alternative splicing that produces a truncated, unstable SMN protein. A small fraction of the SMN2 transcripts include exon 7 and encode the same SMN protein as SMN1. SMA severity correlates with levels of the SMN protein, with those infants with low copy number of SMN2 genes dying within two years. Children with milder forms of SMA have multiple copies of SMN2. This dose-dependency creates the opportunity for treatment of SMA by up-regulating expression of the full-length SMN protein from SMN2. We implemented a lead discovery program to identify drug-like compounds that increase SMN protein levels. Using a novel cell-based reporter assay, we performed high-throughput screens (HTS) of large chemically diverse libraries. We have validated the activities of multiple hits in secondary tests for SMN protein expression and are presently pursuing the optimization of three chemically distinct scaffolds through a focused medicinal chemistry program. Aim 1 will test the drug-like properties, pharmacokinetics and acute toxicology of active compounds that emerge from these structure-activity relationship studies. Aim 2 will examine the most active and pharmacologically suitable compounds in the "7" SMA mouse model to determine whether compound treatment increases SMN protein in vivo and whether the disease process can be mitigated. Aim 3 describes a combinatorial approach to examine the efficacy of the most promising compounds. Since compounds identified in the cell-based HTS appear to act through different mechanisms to stimulate SMN levels, a treatment approach to maximize SMN induction and minimize toxicity will be undertaken by evaluating the biological activity of pair-wise combinations of these leads. This proposal merges the efforts of three laboratories with expertise in SMA molecular biology, development of novel therapies for neurodegenerative diseases, and performance of drug trials in murine models of SMA. The predicted outcome of this proposal is identification of pharmacologically suitable compounds that increase SMN levels and restore motor activity in SMA model mice that can be advanced to human trials. The ultimate goal is to develop an effective drug treatment of spinal muscular atrophy, the leading genetic cause of infant mortality. PUBLIC HEALTH RELEVANCE: Spinal muscular atrophy (SMA), a form of muscular dystrophy, is the leading genetic cause of infant death, and in less devastating forms leads to muscle weakness in children and adults. All forms of SMA result from insufficient levels of the protein called SMN. There is no treatment for SMA. We designed a strategy is to individually test very large collections of compounds for ability to specifically increase SMN protein levels. This grant application requests funds for characterizing these chemicals'drug-like properties and testing their efficacy in a mouse model of SMA. Success would engender their optimization followed by entry into human clinical trials.